Browsing by Author "Acuna Castillo, Claudio"
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- ItemReactive Oxygen Species Potentiate the P2X(2) Receptor Activity through Intracellular Cys(430)(SOC NEUROSCIENCE, 2009) Coddou, Claudio; Codocedo, Juan F.; Li, Shuo; Lillo, Juan G.; Acuna Castillo, Claudio; Bull, Paulina; Stojilkovic, Stanko S.; Huidobro Toro, J. PabloP2X receptor channels (P2XRs) are allosterically modulated by several compounds, mainly acting at the ectodomain of the receptor. Like copper, mercury, a metal that induces oxidative stress in cells, also stimulates the activity of P2X(2)R and inhibits the activity of P2X(4)R. However, the mercury modulation is not related to the extracellular residues critical for copper modulation. To identify the site(s) for mercury action, we generated two chimeras using the full size P2X(2) subunit, termed P2X(2a), and a splice variant lacking a 69 residue segment in the C terminal, termed P2X(2b), as the donors for intracellular and transmembrane segments and the P2X(4) subunit as the donor for ectodomain segment of chimeras. The potentiating effect of mercury on ATP-induced current was preserved in Xenopus oocytes expressing P2X(4/2a) chimera but was absent in oocytes expressing P2X(4/2b) chimera. Site-directed mutagenesis experiments revealed that the Cys(430) residue mediates effects of mercury on the P2X(2a)R activity. Because mercury could act as an oxidative stress inducer, we also tested whether hydrogen peroxide (H2O2) and mitochondrial stress inducers myxothiazol and rotenone mimicked mercury effects. These experiments, done in both oocytes and human embryonic kidney HEK293 cells, revealed that these compounds potentiated the ATP-evoked P2X(2a)R and P2X(4/2a)R currents but not P2X(2b)R and P2X(2a)-C430A and P2X(2a)-C430S mutant currents, whereas antioxidants dithiothreitrol and N-acetylcysteine prevented the H2O2 potentiation. Alkylation of Cys(430) residue with methylmethane-thiosulfonate also abolished the mercury and H2O2 potentiation. Altogether, these results are consistent with the hypothesis that the Cys(430) residue is an intracellular P2X(2a)R redox sensor.
- ItemThe release of sympathetic neurotransmitters is impaired in aged rats after an inflammatory stimulus: A possible link between cytokine production and sympathetic transmission(ELSEVIER IRELAND LTD, 2008) Donoso, Veronica; Gomez, Christian R.; Orriantia, Miguel Angel; Perez, Viviana; Torres, Claudio; Coddou, Claudio; Nelson, Pablo; Maisey, Kevin; Morales, Bernardo; Fernandez, Ricardo; Imarai, Monica; Huidobro Toro, Juan Pablo; Sierra, Felipe; Acuna Castillo, ClaudioAging results in a general decline in the response to external insults, including acute inflammatory challenges. In young animals, the inflammatory response requires activation of the sympathetic system, including neurotransmitters Such as ATP, and catecholamines (epinephrine and norepinephrine). To test whether aging affects activation of this axis, and whether this in turn might affect cytokine release, we administered lipopolysaccharide (LPS) i.p. to adult, middle-aged and aged Fisher 344 rats (6-, 15- and 23-month old, respectively) and evaluated the early (0-12 h) serum levels of Neuropeptide-Y (NP-Y), ATP and vanillyl mandelic acid (VMA, as in indirect measurement of catecholamine levels). In addition, we evaluated the association between these factors and serum levels of the cytokines tumor necrosis factor-alpha (TNF alpha) and interleukin-10 (IL-10). Induction of both ATP and NP-Y was markedly reduced in the serum of aged animals, when compared to their younger Counterparts, while induction of VMA was not affected by age. in spite of these changes, serum levels of TNF alpha and IL-10 were strongly hyper induced and delayed in aged rats. The results suggest that during aging there is a dys-regulation in sympathetic neurotransmitter regulatory Mechanisms, and this might play a role in the impairment of the inflammatory response. (C) 2008 Elsevier Ireland Ltd. All rights reserved.